Immunological and virological findings in a patient with exceptional post-treatment control: a case report.

BACKGROUND: Although antiretroviral therapy (ART) is effective in suppressing viral replication, HIV-1 persists in reservoirs and rebounds after ART has been stopped. However, a very few people (eg, elite and post-treatment controllers) are able to maintain viral loads below detection limits without ART, constituting a realistic model for long-term HIV remission. Here, we describe the HIV control mechanisms of an individual who showed exceptional post-treatment control for longer than 15 years. METHODS: We report the case of a Hispanic woman aged 59 years with sexually acquired acute HIV infection, who was included in an immune-mediated primary HIV infection trial involving a short course of ciclosporine A, interleukin-2, granulocyte macrophage colony-stimulating factor, and pegylated interferon alfa, followed by analytical treatment interruption. We did the following viral assays: total and integrated HIV-1 DNA in CD4 T cells and rectal tissue, quantitative viral outgrowth assay, HIV-1 infectivity in peripheral blood mononuclear cells and CD4 T-cell cultures and viral inhibitory activity by natural killer (NK) and CD8 T cells. NK and T-cell phenotypes were determined by flow cytometry. HLA, killer cell immunoglobulin-like receptors, Δ32CCR5, and NKG2C alleles were genotyped. FINDINGS: After ART and immunomodulatory treatment, the person maintained undetectable plasma viral load for 15 years. HIV-1 subtype was CFR_02AG, CCR5-tropic. We found progressive reductions in viral reservoir during the 15-year treatment interruption: total HIV DNA (from 4573·50 copies per 106 CD4 T cells to 95·33 copies per 106 CD4 T cells) and integrated DNA (from 85·37 copies per 106 CD4 T cells to 5·25 copies per 106 CD4 T cells). Viral inhibition assays showed strong inhibition of in vitro HIV replication in co-cultures of CD4 T cells with autologous NK or CD8 T cells at 1:2 ratio (75% and 62%, respectively). Co-cultures with NK and CD8 T cells resulted in 93% inhibition. We detected higher-than-reference levels of both NKG2C-memory-like NK cells (46·2%) and NKG2C γδ T cells (64·9%) associated with HIV-1 control. INTERPRETATION: We described long-term remission in a woman aged 59 years who was treated during primary HIV infection and has maintained undetectable viral load for 15 years without ART. Replication-competent HIV-1 was isolated. NKG2C-memory-like NK cells and γδ T cells were associated with the control viral replication. Strategies promoting these cells could bring about long-term HIV remission. FUNDING: Fondo Europeo para el Desarrollo Regional (FEDER), SPANISH AIDS Research Network (RIS), Fondo de Investigación Sanitaria (FIS), HIVACAT, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), CERCA Programme/Generalitat de Catalunya, la Caixa Foundation, and Centro de Investigación Biomédica en Red de Enfermedades Infecciosas (CIBERINFEC). TRANSLATION: For the Spanish translation of the abstract see Supplementary Materials section.

Highly Efficient Autologous HIV-1 Isolation by Coculturing Macrophage With Enriched CD4+ T Cells From HIV-1 Patients.

We described a novel HIV autologous isolation method based in coculturing macrophages and CD4+T-cell-enriched fractions from peripheral blood collected from antiretroviral-treated (ART) HIV patients. This method allows the isolation of high viral titers of autologous viruses, over 1010HIV RNA copies/ml, and reduces the time required to produce necessary amounts for virus for use as antigens presented by monocyte-derived myeloid cells in HIV therapeutic vaccine approaches. By applying these high titer and autologous virus produced in the patient-derived cells, we intended to elicit a boost of the immunological system response in HIV therapeutic vaccines in clinical trials.

Interferon gamma transcript detection on T cells based on magnetic actuation and multiplex double-tagging electrochemical genosensing.

Interferon-γ is a proinflammatory cytokine, and its production is related with effective host defense against intracellular pathogens. Therefore, the level of interferon-γ is considered a good biomarker for intracellular infections. It is also useful for the assessment, treatment progression and follow-up of non-communicable diseases, including cancer and autoimmune disorders, among others. This work addresses the development of a novel interferon-γ release assay (IGRA) to evaluate the expression of interferon-γ transcripts as biomarker produced by isolated T cells, as a main advantage. The method sequentially combined three different types of magnetic separation, including the immunomagnetic separation of the T cells performed on antiCD3 modified magnetic particles, the retrotranscription and multiplex double-tagging PCR on polydT-modified magnetic particles and, finally, the electrochemical genosensing on streptavidin magnetic particles as a support. This approach is able to quantify the levels of cellular interferon-γ produced by as low as 150 T cells with outstanding analytical features. The detection of interferon-γ transcripts is performed from only 100 µL of whole blood which can be potentially obtained by fingerprick, demonstrating a further clear advantage to be considered as a promising strategy for the quantification of this important biomarker in several clinical applications.

Interleukin-13 Pathway Alterations Impair Invariant Natural Killer T-Cell-Mediated Regulation of Effector T Cells in Type 1 Diabetes.

Many studies have shown that human natural killer T (NKT) cells can promote immunity to pathogens, but their regulatory function is still being investigated. Invariant NKT (iNKT) cells have been shown to be effective in preventing type 1 diabetes in the NOD mouse model. Activation of plasmacytoid dendritic cells, modulation of B-cell responses, and immune deviation were proposed to be responsible for the suppressive effect of iNKT cells. We studied the regulatory capacity of human iNKT cells from control subjects and patients with type 1 diabetes (T1D) at disease clinical onset. We demonstrate that control iNKT cells suppress the proliferation of effector T cells (Teffs) through a cell contact-independent mechanism. Of note, suppression depended on the secretion of interleukin-13 (IL-13) by iNKT cells because an antibody blocking this cytokine resulted from the abrogation of Teff suppression; however, T1D-derived iNKT cells showed impaired regulation that could be attributed to the decrease in IL-13 secretion. Thus, alteration of the IL-13 pathway at disease onset may lead to the progression of the autoimmune response in T1D. Advances in the study of iNKT cells and the selection of agonists potentiating IL-13 secretion should permit new therapeutic strategies to prevent the development of T1D.

Low frequency of GITR+ T cells in ex vivo and in vitro expanded Treg cells from type 1 diabetic patients.

Reported alterations in T(reg) cells from type 1 diabetes (T1D) patients led us to a revision of their phenotypical features compared with controls. A fine cytometric analysis was designed for their characterization, using a panel of markers including FOXP3, CTLA4, glucocorticoid-induced TNFR family related (GITR) and CD127. The frequency of peripheral CD4(+)CD25(hi) T(reg) cells was similar between samples. However, the yield of sorted T(reg) cells was significantly lower in patients than in controls. When comparing the T(reg)-cell phenotype between samples, the only difference concerned the expression of GITR. A significant decrease of GITR(+) cells and GITR mean fluorescence intensity within the T(reg)-cell population, and to a lesser extent in the effector population, was observed in T1D compared with controls. Moreover, GITR expression was analyzed in several conditions of T-cell activation and differences were only observed in T1D T(reg) cells versus controls when responding to sub-optimal stimulation, that is, soluble anti-CD3 or medium alone but not in the presence of anti-CD3-/anti-CD28-coated beads. However, expanded T1D T(reg)-cell-mediated suppression was as efficient as that mediated by their control counterparts, showing no association between their regulatory capacity and the reduced GITR. Our results show a higher susceptibility to apoptosis in patients' versus controls' T(reg) cells, suggesting that GITR is a T(reg)-cell marker that would be primarily involved in T(reg)-cell survival rather than in their suppressor function.

TCR bias of in vivo expanded T cells in pancreatic islets and spleen at the onset in human type 1 diabetes.

Autoreactive T cells, responsible for the destruction of pancreatic ß cells in type 1 diabetes, are known to have a skewed TCR repertoire in the NOD mouse. To define the autoreactive T cell repertoire in human diabetes, we searched for intraislet monoclonal expansions from a recent onset in human pancreas to then trace them down to the patient's peripheral blood and spleen. Islet infiltration was diverse, but five monoclonal TCR ß-chain variable expansions were detected for Vß1, Vß7, Vß11, Vß17, and Vß22 families. To identify any sequence bias in the TCRs from intrapancreatic T cells, we analyzed 139 different CDR3 sequences. We observed amino acid preferences in the NDN region that suggested a skewed TCR repertoire within infiltrating T cells. The monoclonal expanded TCR sequences contained amino acid combinations that fit the observed bias. Using these CDR3 sequences as a marker, we traced some of these expansions in the spleen. There, we identified a Vß22 monoclonal expansion with identical CDR3 sequence to that found in the islets within a polyclonal TCR ß-chain variable repertoire. The same Vß22 TCR was detected in the patient's PBMCs, making a cross talk between the pancreas and spleen that was reflected in peripheral blood evident. No other pancreatic monoclonal expansions were found in peripheral blood or the spleen, suggesting that the Vß22 clone may have expanded or accumulated in situ by an autoantigen present in both the spleen and pancreas. Thus, the patient's spleen might be contributing to disease perpetuation by expanding or retaining some autoreactive T cells.